HPLC -problem with solvent peak - (Jul/20/2009 )

i got problem with HPLC.
my sample was dissolved in hexane and this solvent was came out in the middle of those compound.
is that possible the solvent peak come out in the middle of analayze compound?
but the peak separation is okay. the only problem is the solvent peak!

thank you very much

-mimosa-

hello Mimosa,
what i know is that the solvent peak appears at the beginning of the run...
what are the remaining compoonds ???
and what's the coloumn used ???

-nightingale-

mimosa on Jul 21 2009, 02:36 AM said:

Hi,

i got problem with HPLC.
my sample was dissolved in hexane and this solvent was came out in the middle of those compound.
is that possible the solvent peak come out in the middle of analayze compound?
but the peak separation is okay. the only problem is the solvent peak!

thank you very much

Try injecting solvent alone and see where it comes.

-swanny-

Hey,

I doubt if its the solvent peak. Did you wash the column nicely before using...maybe some left over by the previous user or some reaction product >. However, running the colvent alone as Swanny suggests will resolve the issue.

Best,
TC

mimosa on Jul 20 2009, 11:06 PM said:

Hi,

i got problem with HPLC.
my sample was dissolved in hexane and this solvent was came out in the middle of those compound.
is that possible the solvent peak come out in the middle of analayze compound?
but the peak separation is okay. the only problem is the solvent peak!

thank you very much

-T C-

thank you for u all suggestion.

i injected and confirm that peak was solvent peak.
i used hexane to dissolve sample and the mobile phase is methanol.
i figure out that hexane not miscible with methanol.
i wondering that the solvent peak in middle of those compound will affect the quantitation?

thank you very much

-mimosa-

mimosa on Jul 21 2009, 10:56 AM said:

i wondering that the solvent peak in middle of those compound will affect the quantitation?

it probably will. integration software will "solvent skim" to try to offset the effect of a solvent peak but the solvent peak and the sample peak shape will have to be consistent from one run to the next.

-mdfenko-

Is there anything stopping you drying off the hexane and redissolving your sample in methanol (or at least something more compatible with your mobile phase)?

-DRT-

why the solvent peak in middle will affect the quantitation?
is that better the solvent peak elute at the beginning?

i try to dissolve the sample in methanol.
but the sample not totally dissolve because i could see the fat globule.

thank you

-mimosa-

mimosa on Jul 22 2009, 09:01 AM said:

why the solvent peak in middle will affect the quantitation?
is that better the solvent peak elute at the beginning?

i try to dissolve the sample in methanol.
but the sample not totally dissolve because i could see the fat globule.

thank you

sorry, i thought you meant that the solvent peak came out with the other components (rising from the solvent peak). if the solvent peak is discrete then your quantitation will be unaffected (unless you are looking at percent of all peaks eluted, then i would turn off integration during the solvent elution).

what would worry me is that the sample is not completely solubilized by the mobile phase (methanol). i would expect the fat globules to form in the column as they separate from the hexane during the run.

can you run the analysis in hexane instead of methanol or, at least, add hexane to the mobile phase?

-mdfenko-

hexane can use as mobile phase for reverse phase column?
because i using c8 column